Calcium entry into stereocilia drives adaptation of the mechanoelectrical transducer current of mammalian cochlear hair cells

Mechanotransduction in the auditory and vestibular systems depends on mechanosensitive ion channels in the stereociliary bundles that project from the apical surface of the sensory hair cells. In lower vertebrates, when the mechanoelectrical transducer (MET) channels are opened by movement of the bundle in the excitatory direction, Ca2+ entry through the open MET channels causes adaptation, rapidly reducing their open probability and resetting their operating range. It remains uncertain whether such Ca2+-dependent adaptation is also present in mammalian hair cells. Hair bundles of both outer and inner hair cells from mice were deflected by using sinewave or step mechanical stimuli applied using a piezo-driven fluid jet. We found that when cochlear hair cells were depolarized near the Ca2+ reversal potential or their hair bundles were exposed to the in vivo endolymphatic Ca2+ concentration (40 µM), all manifestations of adaptation, including the rapid decline of the MET current and the reduction of the available resting MET current, were abolished. MET channel adaptation was also reduced or removed when the intracellular Ca2+ buffer 1,2-Bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid (BAPTA) was increased from a concentration of 0.1 to 10 mM. The findings show that MET current adaptation in mouse auditory hair cells is modulated similarly by extracellular Ca2+, intracellular Ca2+ buffering, and membrane potential, by their common effect on intracellular free Ca2+. Hearing and balance depend on the transduction of mechanical stimuli into electrical signals. This process depends on the opening of mechanoelectrical transducer (MET) channels located at the tips of the shorter of pairs of adjacent stereocilia (1), which are specialized microvilli-like structures that form the hair bundles that project from the upper surface of hair cells (2,3). Deflection of hair bundles in the excitatory direction (i.e., toward the taller stereocilia) stretches specialized linkages, the tip-links, present between adjacent stereocilia (3⇓–5), opening the MET channels. In hair cells from lower vertebrates, open MET channels reclose during constant stimuli via an initial fast adaptation mechanism followed by a much slower, myosin-based motor process, both of which are driven by Ca2+ entry through the channel itself (6⇓⇓⇓⇓⇓⇓–13). In mammalian auditory hair cells, MET current adaptation seems to be mainly driven by the fast mechanism (14⇓–16), although the exact process by which it occurs is still largely unknown. The submillisecond speed associated with the adaptation kinetics of the MET channels in rat and mouse cochlear hair cells (17, 18) indicates that Ca2+, to cause adaptation, has to interact directly with a binding site on the channel or via an accessory protein (16). However, a recent investigation on rat auditory hair cells has challenged the view that Ca2+ entry is required for fast adaptation, and instead proposed an as-yet-undefined mechanism involving a Ca2+-independent reduction in the viscoelastic force of elements in series with the MET channels (19). In the present study, we further investigated the role of Ca2+ in MET channel adaptation in mouse cochlear hair cells by deflecting their hair bundles using a piezo-driven fluid jet, which is believed to produce a more uniform deflection of the hair bundles (20⇓⇓–23) compared with the piezo-driven glass rod (19, 24)

Tmc1 point mutation affects Ca2+ sensitivity and block by dihydrostreptomycin of the mechanoelectrical transducer current of mouse outer hair cells

The transduction of sound into electrical signals depends on mechanically sensitive ion channels in the stereociliary bundle. The molecular composition of this mechanoelectrical transducer (MET) channel is not yet known. Transmembrane channel-like protein isoforms 1 (TMC1) and 2 (TMC2) have been proposed to form part of the MET channel, although their exact roles are still unclear. Using Beethoven (Tmc1Bth/Bth) mice, which have an M412K point mutation in TMC1 that adds a positive charge, we found that Ca2+ permeability and conductance of the MET channel of outer hair cells (OHCs) were reduced. Tmc1Bth/Bth OHCs were also less sensitive to block by the permeant MET channel blocker dihydrostreptomycin, whether applied extracellularly or intracellularly. These findings suggest that the amino acid that is mutated in Bth is situated at or near the negatively charged binding site for dihydrostreptomycin within the permeation pore of the channel. We also found that the Ca2+ dependence of the operating range of the MET channel was altered by the M412K mutation. Depolarization did not increase the resting open probability of the MET current of Tmc1Bth/Bth OHCs, whereas raising the intracellular concentration of the Ca2+ chelator BAPTA caused smaller increases in resting open probability in Bth mutant OHCs than in wild-type control cells. We propose that these observations can be explained by the reduced Ca2+ permeability of the mutated MET channel indirectly causing the Ca2+ sensor for adaptation, at or near the intracellular face of the MET channel, to become more sensitive to Ca2+ influx as a compensatory mechanism

Composite Overwrapped Pressure Vessels (COPV): Flight Rationale for the Space Shuttle Program

Each Orbiter Vehicle (Space Shuttle Program) contains up to 24 Kevlar49/Epoxy Composite Overwrapped Pressure Vessels (COPV) for storage of pressurized gases. In the wake of the Columbia accident and the ensuing Return To Flight (RTF) activities, Orbiter engineers reexamined COPV flight certification. The original COPV design calculations were updated to include recently declassified Kevlar COPV test data from Lawrence Livermore National Laboratory (LLNL) and to incorporate changes in how the Space Shuttle was operated as opposed to orinigially envisioned. 2005 estimates for the probability of a catastrophic failure over the life of the program (from STS-1 through STS-107) were one-in-five. To address this unacceptable risk, the Orbiter Project Office (OPO) initiated a comprehensive investigation to understand and mitigate this risk. First, the team considered and eventually deemed unfeasible procuring and replacing all existing flight COPVs. OPO replaced the two vessels with the highest risk with existing flight spare units. Second, OPO instituted operational improvements in ground procedures to signficiantly reduce risk, without adversely affecting Shuttle capability. Third, OPO developed a comprehensive model to quantify the likelihood of occurrance. A fully-instrumented burst test (recording a lower burst pressure than expected) on a flight-certified vessel provided critical understanding of the behavior of Orbiter COPVs. A more accurate model was based on a newly-compiled comprehensive database of Kevlar data from LLNL and elsewhere. Considering hardware changes, operational improvements and reliability model refinements, the mean reliability was determined to be 0.998 for the remainder of the Shuttle Program (from 2007, for STS- 118 thru STS-135). Since limited hardware resources precluded full model validation through multiple tests, additional model confidence was sought through the first-ever Accelerated Stress Rupture Test (ASRT) of a flown flight article. A Bayesian statistical approach was developed to interpret possible test results. Since the lifetime observed in the ASRT exceeded initial estimates by one to two orders of magnitude, the Space Shuttle Program deemed there was significant conservatism in the model and accepted continued operation with existing flight hardware. Given the variability in tank-to-tank original prooftest response, a non-destructive evaluation (NDE) technique utilizing Raman Spectroscopy was developed to directly measure COPV residual stress state. Preliminary results showed that patterns of low fiber elastic strains over the outside vessel surface, together with measured permanent volume growth during proof, could be directly correlated to increased fiber stress ratios on the inside fibers adjacent to the liner, and thus reduced reliability

Holonomy on D-Branes

This paper shows how to construct anomaly free world sheet actions in string theory with $D$-branes. Our method is to use Deligne cohomology and bundle gerbe theory to define geometric objects which are naturally associated to $D$-branes and connections on them. The holonomy of these connections can be used to cancel global anomalies in the world sheet action.Comment: Corrections made and some typographical errors remove

Sorghum: An Underutilized Cereal Whole Grain with the Potential to Assist in the Prevention of Chronic Disease

Sorghum is an important cereal grain food, grown globally, that is rich in nutrients, dietary fiber, and bioactive components yet is considered of low value to humans and often used as an animal feed. This review provides an overview of key sorghum grain components, including starches, dietary fiber, protein, lipids, and phytochemicals, with functional properties that have potential to impact on health. Though acknowledging the impact of the whole food will reflect the synergy between the components, studies of these components implicate effects on energy balance, glycemic control, lipids, gut microbiota, and cell-mediated immune responses, including antioxidant and anti-inflammatory effects. For these to be confirmed as contributory effects from sorghum consumption, evidence from quality randomized controlled trials is required. If proven effective, there may be a role for sorghum grain–based diets to assist in the prevention of chronic diseases such as diabetes, obesity, and heart disease. Future research addressing effects of sorghum consumption may help drive a paradigm shift from sorghum as a low value food to a potentially health-promoting, highly valued human grain food

Hair cell maturation is differentially regulated along the tonotopic axis of the mammalian cochlea

Sound amplification within the mammalian cochlea depends upon specialized hair cells, the outer hair cells (OHCs), which possess both sensory and motile capabilities. In various altricial rodents, OHCs become functionally competent from around postnatal day 7 (P7), before the primary sensory inner hair cells (IHCs), which become competent at about the onset of hearing (P12). The mechanisms responsible for the maturation of OHCs and their synaptic specialization remain poorly understood. We report that spontaneous Ca2+ activity in the immature cochlea, which is generated by CaV1.3 Ca2+ channels, differentially regulates the maturation of hair cells along the cochlea. Under near‐physiological recording conditions we found that, similar to IHCs, immature OHCs elicited spontaneous Ca2+ action potentials (APs), but only during the first few postnatal days. Genetic ablation of these APs in vivo, using CaV1.3−/− mice, prevented the normal developmental acquisition of mature‐like basolateral membrane currents in low‐frequency (apical) hair cells, such as IK,n (carried by KCNQ4 channels), ISK2 and IACh (α9α10nAChRs) in OHCs and IK,n and IK,f (BK channels) in IHCs. Electromotility and prestin expression in OHCs were normal in CaV1.3−/− mice. The maturation of high‐frequency (basal) hair cells was also affected in CaV1.3−/− mice, but to a much lesser extent than apical cells. However, a characteristic feature in CaV1.3−/− mice was the reduced hair cell size irrespective of their cochlear location. We conclude that the development of low‐ and high‐frequency hair cells is differentially regulated during development, with apical cells being more strongly dependent on experience‐independent Ca2+ APs

Ecophysiology of coral reef primary producers across an upwelling gradient in the tropical central Pacific

© The Author(s), 2020. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Johnson, M. D., Fox, M. D., Kelly, E. L. A., Zgliczynski, B. J., Sandin, S. A., & Smith, J. E. Ecophysiology of coral reef primary producers across an upwelling gradient in the tropical central Pacific. Plos One, 15(2), (2020): e0228448, doi:10.1371/journal.pone.0228448.Upwelling is an important source of inorganic nutrients in marine systems, yet little is known about how gradients in upwelling affect primary producers on coral reefs. The Southern Line Islands span a natural gradient of inorganic nutrient concentrations across the equatorial upwelling region in the central Pacific. We used this gradient to test the hypothesis that benthic autotroph ecophysiology is enhanced on nutrient-enriched reefs. We measured metabolism and photophysiology of common benthic taxa, including the algae Porolithon, Avrainvillea, and Halimeda, and the corals Pocillopora and Montipora. We found that temperature (27.2–28.7°C) was inversely related to dissolved inorganic nitrogen (0.46–4.63 μM) and surface chlorophyll a concentrations (0.108–0.147 mg m-3), which increased near the equator. Contrary to our prediction, ecophysiology did not consistently track these patterns in all taxa. Though metabolic rates were generally variable, Porolithon and Avrainvillea photosynthesis was highest at the most productive and equatorial island (northernmost). Porolithon photosynthetic rates also generally increased with proximity to the equator. Photophysiology (maximum quantum yield) increased near the equator and was highest at northern islands in all taxa. Photosynthetic pigments also were variable, but chlorophyll a and carotenoids in Avrainvillea and Montipora were highest at the northern islands. Phycobilin pigments of Porolithon responded most consistently across the upwelling gradient, with higher phycoerythrin concentrations closer to the equator. Our findings demonstrate that the effects of in situ nutrient enrichment on benthic autotrophs may be more complex than laboratory experiments indicate. While upwelling is an important feature in some reef ecosystems, ancillary factors may regulate the associated consequences of nutrient enrichment on benthic reef organisms.This work was supported by funding from the Moore Family Foundation, the Gordon and Betty Moore Foundation, the Scripps family, and anonymous donors. The funders had no role in study design, data collection and analysis, or preparation of the manuscript

Gold(I)-Catalysed Direct Thioetherifications Using Allylic Alcohols: an Experimental and Computational Study

A gold(I)-catalysed direct thioetherification reaction between allylic alcohols and thiols is presented. The reaction is generally highly regioselective (S(N)2′). This dehydrative allylation procedure is very mild and atom economical, producing only water as the by-product and avoiding any unnecessary waste/steps associated with installing a leaving or activating group on the substrate. Computational studies are presented to gain insight into the mechanism of the reaction. Calculations indicate that the regioselectivity is under equilibrium control and is ultimately dictated by the thermodynamic stability of the products